We developed a method to induce rapid assembly of gap junctions in prostate epithelial cells in vitro. Assembly of gap junctions can be induced by incubation at O degree C followed by incubation at 37 degrees C. Colchicine (10 to the -5 M, 10 to the -3 M) and cytochalasin B (25 Mug/m1, 100 Mug/m1) at room temperature or at 37C also induce assembly of gap junctions. Assembly of the junctions proceeds even in the presence of a metabolic inhibitor (dinitrophenol) or of an inhibitor of protein synthesis (cycloheximide). We conclude that assembly of gap junctions can proceed from a pool of pre-existing precursors. The experimental conditions that result in gap-junction assembly involve perturbation of the cytoskeleton. Therefore, we propose that the assembly of gap junctions requires convergent migration of precursor molecules whose positional control in the membrane is released by perturbation of the cytoskeleton. Aggregates of particles and rugosities, whose distribution size and shape is similar to that of gap junctions, may represent intermediate assembly stages. This would indicate that the final stages in the assembly take place only after convergence of the precursor molecules to the junctional site and involve profound conformation changes required for establishment of fully assembled connexons.